Open AccessCONSTRUCTION OF YEAST PICHIA PASTORIS EXPRESSING SECRETED MINIPROINSULIN INTO CULTURE
Author(s) -
Hang Ngo,
Tri Minh Vo,
Thuoc Linh Tran
Publication year - 2012
Publication title -
khoa học công nghệ
Language(s) - English
Resource type - Journals
ISSN - 1859-0128
DOI - 10.32508/stdj.v15i2.1800
Subject(s) - pichia pastoris , recombinant dna , fusion protein , yeast , plasmid , pichia , gene , fusion gene , biology , microbiology and biotechnology , chemistry , genetics
Diabetes is one of the diseases, which have recently gained significant attention. In order to produce the recombinant insulin for treating diabetics, we have cloned and expressed a miniproinsulin in Pichia pastoris. The DNA fragment containing a gene encoding the fusion protein of miniproinsulin-6xhis was cloned into the integrative plasmid pPICZαA resulting in pPICZαA/h-mpi to construct the strain P. pastoris GS115::h-mpi expressing the recombinant MPI as secreted protein into culture. Phenotype and genotype of P. pastoris GS115::h-mpi were confirmed by PCR and restriction pattern. MPI was expressed as a fusion protein with 6xhis tag by supplementing methanol at a final concentration of 0.5% in BMMY culture medium for 96 hours after inducing.